Modulation of excitation-contraction coupling by isoproterenol in cardiomyocytes with controlled SR Ca2+ load and Ca2+ current trigger

Cardiac Ca2+ transients are enhanced by cAMP-dependent protein kinase (PKA). However, PKA-dependent modulation of ryanodine receptor (RyR) function in intact cells is difficult to measure, because PKA simultaneously increases Ca2+ current (I-Ca), SR Ca2+ uptake and SR Ca2+ loading (which independently increase SR Ca2+ release). We measured Ica and SR Ca2+ release +/- 1 mum isoproterenol (ISO; isoprenaline) in voltage-clamped ventricular myocytes of rabbits and transgenic mice (expressing only non-phosphorylatable phospholamban). This mouse model helps control for any effect of ISO-enhanced SR uptake on observed release, but the two species produced essentially identical results. SR Ca2+ load and Ica were adjusted by conditioning. We thus evaluated PKA effects on SR Ca2+ release at constant SR Ca2+ load and I-Ca trigger (with constant unitary I-Ca). The amount of SR Ca2+ release increased as a function of either Ica or SR Ca2+ load, but ISO did not alter the relationships (measured as gain or fractional release). This was true over a wide range of SR Ca2+ load and Ica. However, the maximal rate of SR Ca2+ release was similar to50% faster with ISO (at most loads and I-Ca levels). We conclude that the isolated effect of PKA on SR Ca2+ release is an increase in maximal rate of release and faster turn-off of release (such that integrated SR Ca2+ release is unchanged). The increased amount of SR Ca2+ release normally seen with ISO depends primarily on increased I-Ca trigger and SR Ca2+ load, whereas faster release kinetics may be the main result of RyR phosphorylation.